Loading Concentration Optimization Recommendations for the MiSeq i100 Series

Background

While many libraries will perform well when loaded in the 80-120 pM range, users running third party or custom libraries will need to perform titration experiments to optimize the loading concentration for these libraries. See the instructions below for recommended started concentrations for specific Illumina and validated third party libraries.

For Third Party libraries not listed or custom libraries, see section Migrating Custom or Third Party Libraries from MiSeq to the MiSeq i100 below for initial guidance.

For additional details on preparing titration experiments, see the section Determining optimal library loading concentration in the Maximizing performance on the MiSeq i100 Series Technical note.

Migrating Custom or Third Party Libraries from MiSeq to the MiSeq i100

When migrating libraries that were optimized for the MiSeq v2 and v3 reagent kits, use the following multipliers to calculate the recommended starting concentration for loading optimization.

MiSeq Reagent Kit	Multiplier
v2 Reagent Kits	~10.4X
v3 Reagent Kits	~6.5X

For example, if the optimized library loading concentration for a customer or third party library is 9 pM using MiSeq v3 reagent kits, the recommended starting concentration for the MiSeq i100 will be ~60 pM. On run completion, titrate the loading concentration up or down in 20 pM increments based on run performance at this starting concentration.

Recommended Loading Concentrations for Validated Illumina and Third Party Libraries

For additional details on recommended loading concentrations, see the Maximizing performance on the MiSeq i100 Series Technical note.

For any feedback or questions regarding this article (Illumina Knowledge Article #9340), contact Illumina Technical Support [e-mail].